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Nucleosome profiles in strains with different inter-NDR distances at HML

ABSTRACT: Heterochromatin is a complex structure comprised of nucleosome arrays that are bound by silencing factors. This composition suggests that certain configurations of nucleosome arrays are permissible for heterochromatic silencing. We tested this idea in S. cerevisiae by systematically altering the distance between heterochromatic Nucleosome Depleted Regions (NDRs), which is predicted to affect local nucleosome positioning by limiting how nucleosomes can be packed between NDRs. Consistent with this prediction, some inter-NDR distances led to poorly positioned nucleosome arrays within heterochromatin. Furthermore, these conditions that caused poor nucleosome positioning also led to defects in both heterochromatin stability and the ability of cells to generate and inherit a silenced transcriptional state. These findings strongly suggest that nucleosome positioning contributes to formation and maintenance of functional heterochromatin.

ORGANISM(S): Saccharomyces cerevisiae

PROVIDER: GSE144808 | GEO | 2020/02/06

REPOSITORIES: GEO

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Project description:Abstract: In the yeast S. pombe, multiple chromatin-modifying enzymes are required for heterochromatin formation, yet how their actions alter chromatin structure to block access to DNAby the transcriptional machinery is unknown. We constructed high-resolution nucleosome occupancy maps of heterochromatic regions in wild-type strains and in mutants lacking the H3K9 methyltransferase Clr4 or one of the two activities of the silencing effector complex SHREC. Fourier analysis reveals that these enzymes do not increase the regularity of nucleosome spacing. Rather, their principal effect was to induce the elimination of nucleosome-free regions (NFRs). Both NFRs associated with transcription initiation sites as well as those not associated with promoters are affected. As in S. cerevisiae, repressed genes in euchromatin retain their NFRs. Thus, NFR elimination cannot be explained as a secondary consequence of repression. The maps also show that TFIIIC boundary elements have NFRs resistant to silencing, suggesting a potential role in preventing lateral spread of heterochromatin. NFR elimination offers a mechanism by which heterochromatin restricts access of the transcriptional machinery to DNA. Nucleosome positions in either heterochromatin or euchromatin were mapped in various mutants to determine whether chromatin structure is altered upon heterochromatic silencing. Eight strains were assayed for nucleosome positioning in heterochromatin while two strains were assayed for nucleosome positioning in euchromatin. Two biological replicates were assayed for each strain. No dye swaps were done. RNA transcripts were also mapped using the heterochromatin tiling arrays to determine where RNA transcripts accumulate in heterochromatin. Three strains were assayed for transcripts in heterochromatin by hybridizing cy5-labeled cDNA created from total RNA against cy3-labeled cDNA created from RNA synthesized from sonicated genomic DNA. Two biological replicates were assayed for each strain.

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Nucleosome profiles in strains with different inter-NDR distances at HML

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